One of the most important goals of dentistry and the NIDCR is to generate a whole Bio-Tooth, which would be composed of crown and root. In the last three decades, considerable achievements have been made in studies of tooth germ and crown formation, although the root studies lag behind due to the lack of appropriate animal models available, plus the inherent difficulties associated with handling mineralized dentin. Nevertheless, it is important to recognize that the root is a critical structural component of the tooth because: i) There are numerous hereditary syndromes and chromosomal anomalies that affect the root structure; and ii) There is an urgent need to generate a root for treatment of dental trauma, tooth agenesis, and periodontal diseases. Our long-term goal is to understand the biological mechanisms controlling root formation, which will fill a key gap of knowledge leading to the goal of regenerating a whole Bio-Tooth. The objective here, which is our next step in the pursuit of that goal, is to define the emerging signaling pathway(s) essential for root but not for crown dentin formation. Our central hypothesis is that the novel control mechanism in root dentin formation is different from that in the tooth crown formation, in which the interaction among NFIC, OSX, -catenin, and DSPP play key roles. This hypothesis is formulated on the basis of our strong preliminary data produced using both global knockout (KO) and conditional KO (cKO) mouse models, as well as in vitro molecular approaches. Two Specific Aims are proposed to test this hypothesis: 1). To demonstrate the critical role of OSX, the key downstream transcriptional factor of NFIC, in control of postnatal root - but not crown - dentin formation by means of the negative regulation of Wnt--catenin and positive regulation of DSPP; and 2).To determine whether blocking the elevated -catenin level rescues the Osx cKO tooth phenotype in vivo and to define whether re- expressions of DSPP either in the Osx cKO or the CA--catenin mice restore dentin tubule formation and mineralization. The proposed research is innovative because i) these genetically engineered animal models developed astonishing tooth root phenotypes with no apparent changes in the crown; and ii) the approaches proposed will delineate the novel roles of each molecule in this emerging signaling pathway in a tissue- and temporal-dependent manner. The studies proposed are significant because the valuable information gained can be applied to fill the critical knowledge gap in this understudied area and contribute to investigations on the generation of a future biological tooth replacement.